Antibody vs Antigen concentration effect
Posted 29 June 2011 - 04:32 AM
Posted 29 June 2011 - 04:49 PM
Posted 30 June 2011 - 05:48 AM
I will explain in more detail. Basically what I am observing is a significant amount of non-specific bands appearing without the band of interest. The amount of lysate loaded onto the gel was 50ug which in our experience should be sufficient. Trying to detect a moderately expressed transcription factor. What was told to me was that if I load more lysate > 100ug that our band of interest will appear and the non-specific bands will disappear. Basically, playing the s/n game. In my experience I have never seen this inversion happen. My guess this only applies to extremely poor, low affinity antibodies. Any insight and/or literature ref explaining this phenomenon would be greatly appreciated.
Posted 30 June 2011 - 05:52 PM
I suppose it could work a bit like PCR - if there is no specific target around, the primers (or antibody) might bind to any sequence and give you some signal.